High-risk human papillomaviruses (HPVs) cause approximately 5% of all human cancers. HPV-associated cancers arise years to decades after the initial infection and hence it will take 20 to 30 years before the available prophylactic vaccines will have an impact on HPV cancer rates. The viral E6 and E7 oncoproteins are necessary for the induction and the maintenance of the transformed state. HPVs contribute to cancer formation by associating with and functionally reprogramming key cellular control circuits. In response, cells attempt to mount innate defense responses to neutralize the invading virus, which the virus has evolved to blunt. To tolerate viral gene expression, host cells have to undergo a variety of adaptive responses. As a consequence, signaling circuits are extensively rewired in HPV infected cells, which renders them uniquely vulnerable to disruption of certain signaling pathways that would be inconsequential in normal, uninfected cells. In the last decade we have discovered multiple cellular vulnerabilities (sometimes referred to as synthetic lethalities) that arise as specific consequences of HPV E6 and/or E7 expression in normal cells. During the previous funding period we discovered that increased p16INK4A expression that is triggered by E7 is mediated by the H3K27 demethylase KDM6B, and represents engagement of cellular defense pathway, referred to as oncogene induced senescence (OIS) that activates the retinoblastoma tumor suppressor, pRB. HPV E7 blunts the OIS response by triggering pRB degradation. Cells continue to proliferate and p16INK4A serves as an excellent biomarker for high-risk HPV associated lesions and cancers. Surprisingly, however, high-risk HPV E7 expressing cells have developed an addiction to KDM6B and p16INK4A and a small molecule KDM6 inhibitor induced apoptosis in E7 expressing cells while not affecting parental cells. This suggests the possibility of epigenetic therapies for HPV associated lesions and cancers. Here we propose to perform a detailed analysis of epigenetic factors that contribute to expression of p16INK4A as well as p14ARF, which is also epigenetically de-repressed by E7 and contributes to a second cellular defense response that triggers p53 tumor suppressor activation. We will also determine whether E7 expressing cells develop addiction to additional epigenetic factors that contribute to p16INK4A expression. Since epigenetic writers, erasers as well as readers are druggable, the insights from these studies may lead to discovery of therapeutic targets for high-risk HPV- associated lesions and cancers.